A method for the isolation of homogeneous fluorescein-labeled proteins for studies on the variation in biological and fluorescence properties of the different conjugates.
Different homogeneous fluorescein-labeled conjugates of alpha-chymotrypsin and the pancreas trypsin inhibitor have been analyzed and isolated by isoelectric focusing with and without biospecific sample application. With this procedure the biological activity of the different protein conjugates can be determined from the focusing pattern. The fluorescence quantum yield was found to be decreased with the number of labels per protein. The biological activity, however, was increased in some monolabeled conjugates compared with the activity of the native protein. The quantum yield was found to be markedly (greater than 10%) changed when the monolabeled conjugates formed specific complexes with proteins.
D000818:Animals; D002417:Cattle; D002918:Chymotrypsin / Q000302:isolation & purification* / Q000378:metabolism; D016650:Fluorescein-5-isothiocyanate; D005452:Fluoresceins / Q000494:pharmacology; D007700:Kinetics; D010179:Pancreas / Q000201:enzymology; D011485:Protein Binding; D013050:Spectrometry, Fluorescence; D013861:Thiocyanates / Q000494:pharmacology; D014357:Trypsin / Q000302:isolation & purification* / Q000378:metabolism
D005452:Fluoresceins; D013861:Thiocyanates; D002918:Chymotrypsin; D014357:Trypsin; D016650:Fluorescein-5-isothiocyanate